Susceptibility to HIV-1 infection varies greatly between individuals, with disease progression rates determined not only by viral properties but also host factors. The central hypothesis of this proposal is that the extensive diversification of the APOBEC3 (A3) family of restriction factors contributes to the high degree of variability in HIV-1 replication in vivo. There is growing evidence that genomic variation - ranging from single nucleotide polymorphisms (SNPs) to large structural variants - influences the efficacy of host antiviral defenses. Indeed, we have evidence that naturally occurring SNPs modulate the antiviral activity of A3F and A3H. Moreover, we have identified three regions near and within the A3 locus itself that show copy number variations among HapMap populations (based on data from 450 individuals of West-African, Asian and European ancestry). These copy number variants (CNV) overlap with A3B, A3E and A3F exons and possibly upstream regulatory sequences of the APOBEC locus. Preliminary data indicate that the APOBEC loci from HIV-1 infected long-term non-progressors (LTNP) are enriched for A3 copy number variants, suggesting a protective role of A3 copy number variation. Conversely, we have shown that cells homozygous for A3-CNV2 copy loss (complete deletion of the A3B gene) are more susceptible to HIV-1 infection in vitro. Collectively, our preliminary data strongly support the notion that A3 regions are structurally polymorphic and that the resulting A3 variants exert a key impact on HIV-1 replication. In specific aim 1 (CNV discovery and mapping), we will interrogate the A3 gene family cluster at very high resolution using cutting edge technologies to capture the wide range of CNVs embedded in this locus. For this purpose, we will use samples from ethnically diverse (18 different groups) or phenotypically selected (LTNP) populations. In specific aim 2 (gene expression), we will determine the mRNA expression pattern specific for A3-CNV genotypes and identify protein variants associated with the panel of rearranged A3 loci. We will also compare the transcriptional regulation of A3 genomic variants in lymphocytes and dendritic cells of LTNPs and treated progressors. In specific aim 3 (gene function), the impact of A3-CNVs and SNPs on viral restriction will be tested, using lymphocytes from individuals with known A3 genotypes, as well as model cell-lines expressing gene products representing naturally circulating A3 genomic variants. These experiments will integrate genomic variation data of the A3 region with an understanding of the regulation and antiviral functions of A3 deaminases. This comprehensive investigation into the functional consequences of the high variability in A3 genomic architecture should help identify new biomarkers to be tested in future epidemiological studies of retroviral infections. In addition, understanding the functional diversity of intrinsic defense factors has the potential to create novel treatment strategies aimed at a long- lasting suppression of HIV/AIDS disease.